Summary Hepatitis B is a global health problem with ~360-400 million chronically infected patients. These individuals can develop fibrosis, cirrhosis and hepatocellular carcinoma resulting frequently in death. A prophylactic vaccine is available and there are anti-HBV drugs that can suppress viremia but there is no cure. The causative agent of hepatitis B is the hepatitis B virus (HBV), a small double stranded DNA virus (3.2 kb genome) with four overlapping open reading frames (ORF's), which produces 4 gene products. The lack of a small animal model for HBV has hampered the study of HBV and the development of more effective therapies. HBV has a narrow host range infecting only humans and chimpanzees. The basis for this highly restricted species tropism is not understood and is the focus of my studies. Previous data demonstrate that while infectious HBV virions assemble and are released from mouse cells, viral entry and genome replication are not supported in this species. It is my goal to systematically dissect blocks in the HBV life-cycle focusing here on HBV uptake in rodent cells. Recently, the human sodium taurocholate co-transporting polypeptide (hNTCP) has been identified as a receptor for HBV and HDV entry. Expression of hNTCP in murine cells will facilitate the uptake of hepatitis delta virus (HDV), a satellite virus that uses the HBV envelope proteins to produce infectious virions, but not HBV. This suggests that there are either additional human host factors besides hNTCP necessary for HBV entry or that there are murine dominant negative factors that restrict HBV uptake. The purpose of this study is to understand the restrictive tropism of HBV and to identify factors that are necessary for entry and to understand their function. The proposed study will result in several significant advancements. First, the development of a novel HBV reporter systems will be created to demonstrate two important principles, 1) their utility as a reporter for HBV entry, and 2) the ability to express heterologous proteins within a HBV virion. Secondly, a genetic screen will be performed to identify additional human host factors besides hNTCP that are necessary for HBV uptake. Collectively, the successful completion of this project will result in novel tools to investigate HBV entry and the identification of human host factors necessary for HBV entry. The identification and characterization of additional human host factors that are important for HBV uptake will form the blue-print to create a mouse model with inheritable susceptibility to HBV infection. This model could be used for testing of novel therapies.